期刊
RAPID COMMUNICATIONS IN MASS SPECTROMETRY
卷 24, 期 15, 页码 2277-2282出版社
WILEY
DOI: 10.1002/rcm.4627
关键词
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资金
- Yamagata Prefecture and Tsuruoka City
- JSPS [21310129]
- Japan Science and Technology Agency
- Grants-in-Aid for Scientific Research [21310129] Funding Source: KAKEN
We have developed a novel strategy to improve the efficiency of identification of multiply phosphorylated peptides isolated by hydroxy acid modified metal oxide chromatography (HAMMOC). This strategy consists of alkali-induced chemical dephosphorylation (beta-elimination reaction) of phosphopeptides isolated by HAMMOC prior to analysis by liquid chromatography/mass spectrometry (LC/MS). This approach identified 1.9-fold more multiply phosphorylated peptides than the conventional approach without beta-elimination from a digested mixture of three standard phosphoproteins. In addition, the accuracy of phosphorylation site determination in synthetic phosphopeptides was significantly improved. Finally, we applied this approach to a cell lysate. By combining this dephosphorylation approach with the conventional approach, we successfully identified 1649 unique phosphopeptides, including 325 multiply phosphorylated phosphopeptides, from 200 mu g of cultured Arabidopsis cells. These results indicate that chemical dephosphorylation prior to LC/MS analysis increases the efficiency of identification of multiply phosphorylated peptides, as well as the accuracy of phosphorylation site determination. Copyright (C) 2010 John Wiley & Sons, Ltd.
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